MONOCLONAL ANTIBODY-BASED ENZYME-IMMUNOASSAY FOR SIMULTANEOUS QUANTITATION OF 2-ALPHA-HYDROXYESTRONE AND 16-ALPHA-HYDROXYESTRONE IN URINE
MONOCLONAL ANTIBODY-BASED ENZYME-IMMUNOASSAY FOR SIMULTANEOUS QUANTITATION OF 2-ALPHA-HYDROXYESTRONE AND 16-ALPHA-HYDROXYESTRONE IN URINE
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DOI：
10.1016/0039-128x(94)90021-3
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发表时间：
1994-11-01
期刊：
影响因子：
2.7
通讯作者：
SEPKOVIC, DW
中科院分区：
文献类型：
--
作者：
KLUG, TL;BRADLOW, HL;SEPKOVIC, DW
雌激素代谢的改变被认为是妇科癌症和红斑狼疮等疾病的病因学中的一个重要因素。雌二醇的主要代谢产物在C-2或C-16 α位羟基化，分别产生具有雌激素拮抗剂和激动剂活性的产物。一个敏感和特异的免疫诊断检测，以确定这些竞争途径之间的平衡可能作为一个常规的生物标志物远雌激素相关疾病的管理。我们在这里描述了高亲和力，特异性鼠单克隆抗体的产生2-羟基雌酮和16 α-羟基雌酮的高效融合协议。利用这些抗体，我们研制了一种快速、简便的酶免疫分析（EIA）试剂盒，用于同时定量未提取尿液中的2-和16 α-羟基雌酮。初步验证研究确定，通过ETA发现的尿代谢物2-和16 α-羟基雌酮浓度与气相色谱-质谱法发现的值相关性良好。EIA试剂盒的初步研究发现，加标尿样中的代谢物总回收率。使用当前EIA试剂盒，2-羟基雌酮和16 α-羟基雌酮的EIA批间和批内变异系数以及2-羟基雌酮与16 α-羟基雌酮的比率始终小于9%。该试剂盒命名为ESTRAMET（（TM））2/16，可能为雌激素相关疾病的研究提供重要的新途径。
Alterations in the metabolism of estrogen hare been implicated as an important factor in the etiology of diseases such as gynecological cancers and lupus erythematosus. The major metabolites of estradiol are hydroxylated at the C-2 or C-16 alpha position yielding products with estrogen antagonist and agonist activities, respectively. A sensitive and specific immunodiagnostic assay to determine the balance between these competing pathways might serve as a routine biomarker far management of estrogen-related diseases. We describe here the generation of high affinity, specific murine monoclonal antibodies to 2-hydroxyesterone and 16 alpha-hydroxyestrone by high efficiency fusion protocols. With these antibodies, we have developed a rapid and simple enzyme immunoassay (EIA) kit for the simultaneous guantitation of 2- and 16 alpha-hydroxyestrone in unextracted urine. Initial validation studies established that urinary metabolite 2- and 16 alpha-hydroxyestrone concentrations found by the ETA correlate well with values found by gas chromatography-mass spectroscopy. Preliminary studies with the EIA kit found total recovery of metabolites from spiked urine samples. The EIA inter- and intra-assay coefficients of variation for 2-hydroxyestrone and 16 alpha-hydroxyestrone and the ratio of 2-hydroxyestrone to 16 alpha-hydroxyestrone with the current EIA kit were consistently less than 9%. This kit, designated ESTRAMET((TM)) 2/16 may provide an important new too[for research in estrogen-related diseases.